This invention relates to combinatorial chemistry libraries containing thiourea and urea compounds. In addition, the invention relates to methods for constructing combinatorial chemistry libraries containing thiourea and urea compounds. Furthermore, this invention relates to methods for the identification of bioactive thiourea and urea compounds as well as compositions and therapeutic methods for treating cancer.
A common method of drug discovery is to first delineate a biochemical pathway that is involved in a targeted pathological process. The biological pathway is analyzed so as to determine crucial elements which, if obstructed, restrained or otherwise adversely modified could inhibit the pathological process. Generally, an assay can be developed that is indicative of the functional ability of an element of the biochemical pathway. The assay can then be performed in the presence of a number of different molecules. The researcher can then determine the molecules that have the desired effect on the pathway, and that molecule or molecules can be used in treatment or can be further modified to augment and enhance the desired effect.
As the assays that are indicative of these pathways become faster, and more easily automated, the rate determining step regarding molecular screening becomes the production of the molecules to be tested. Thus, the development of techniques to rapidly and systematically synthesize large numbers of molecules possessing diverse structural properties has grown in importance. On such technique for rapidly and systematically synthesizing large numbers of molecules possessing diverse structural properties is the construction of combinatorial libraries. Combinatorial chemistry employing solution-phase combinatorial synthesis plays and increasingly important role in drug discovery efforts.
Combinatorial libraries are typically formed via a multistep synthetic procedure employing either solution-phase or solid-phase methods. The procedure typically includes mixtures of different subunits which are added stepwise to growing oligomers until a desired oligomer size is reached. Alternatively, the subunits can be combined in one synthetic step to produce a random array of oligomers or a combination of the two procedures may be employed. The result is the rapid synthesis of a large, diverse group of chemical compounds that can be screened with the predictive assay developed with regard to the targeted pathological process. Since the chance of finding useful molecules increases with the size of the combinatorial library, it is desirable to generate libraries composed of large numbers of oligomers which vary in their subunit sequence.
Apoptosis is a biochemical process that is an important part of a number of diseases. Apoptosis is a common mode of eukaryotic cell death which is triggered by an inducible cascade of biochemical events leading to activation of endonucleases that cleave the nuclear DNA into oligonucleosome-length fragments. Several of the biochemical events that contribute to apoptotic cell death as well as both positive and negative regulators of apoptosis have recently been identified (Whyllie A., et al. (1980) Int. Rev. Cytol. 68, 251-305; Steller H., (1995) Science 267, 1445-1449; Fraser, A., Evan, G. (1996) Cell 85, 781-784; and Korsmeyer, S. J. (1995). Trends Genet. 11, 101-105). Apoptosis plays a pivotal role in the development and maintenance of a functional immune system by ensuring the timely self-destruction of autoreactive immature and mature lymphocytes as well as any emerging target neoplastic cells by cytotoxic T cells.
In addition to the beneficial effects associated with apoptosis, inappropriate apoptosis contributes to the pathogenesis and drug resistance of human leukemias and lymphomas (Cohen, J. J., et al. (1992) Annu. Rev. Immunol. 10, 267-293; Linette, G. P., Korsmeyer, S. J. (1994) Curr. Opin. Cell Biol. 6, 809-815; and Thompson, C. B. (1995) Science 367, 1456-1462). Thus, agents that are useful to modulate apoptosis are potentially useful as therapeutic agents for treating diseases in which inappropriate apoptosis is implicated. As a result, there is a considerable amount of ongoing research devoted to the identification of molecular regulators of apoptosis, and there is currently a need for novel agents (e.g. chemical or biological), and novel therapeutic methods, that are useful for modulating apoptosis. Such agents and methods may be useful for treating cancer (e.g. leukemias and lymphomas) or immune disorders in mammals. They may also be useful as pharmacological tools for use in in vitro or in vivo studies to enhance the understanding of the molecular basis of apoptosis (e.g. the pro-apoptotic versus the anti-apoptotic regulatory signal), as well as the pathogenesis of human lymphoid malignancies.
Novel thiourea and urea compounds have been found to be potent cytotoxic agents with potent activity against cancer cells. For example, certain thiourea and urea compounds exhibit potent cytotoxic activity, particularly against human leukemic cell lines. Additionally, thiourea and urea compounds have been found to be nonnucleoside inhibitors of HIV reverse transcriptase. Currently the production of thiourea and urea compounds however, is limited to the small scale synthesis of individual molecules. Thus, a method for the rapid and systematic synthesis of large numbers of thiourea and urea compounds possessing diverse structural properties is desirable.
Generally, the present invention relates to a combinatorial library including compounds of the Formula I 
wherein X is S or O;
R and R1 are individually 
where Ar is aryl; R2 is H or C1 to C6 alkyl; n is 0-3 and where the aryl moiety is optionally substituted from 1 to 7 times with any combination of H, halo, alkyl, haloalkyl, arylalkyl, alkoxy, haloalkoxy, and aralkoxy. One embodiment is a combinatorial library of claim 1, wherein R and R1 are individually 
where R2 is H or C1 to C6 alkyl; n is 0-3 and where the phenyl moiety is optionally substituted from 1 to 5 times with any combination of R3 R4, R5, R6, R7, R8, R9, R10, R11, R12, R13, R14, R15, R16, R17, R18, R19, R20, R21, R22, R23, R24, R25, R26, and R27; and where R3 is H, R4 is 2-methyl, R5 is 2-trifluoromethyl, R6 is 2-fluoro, R7 2-chloro, R8 is 2-methoxy, R9 is 2-ethoxy, R10 3-methyl, R11 is 3-trifluoromethyl, R12 is 3-fluoro, R13 is 3-chloro, R14 is 3-iodo, R15 is 3-methoxy, R16 is 4-methyl, R17 is 4-trifluoromethyl, R18 is 4-fluoro, R19 is 4-chloro, R20 is 4-bromo, R21 is 4-methoxy, R22 is 5-trifluoromethyl, R23 is 5-fluoro, R24 is 6-fluoro, R25 is 5-methoxy, R26 is 3-benzyloxy, and R27 is 4-benzyloxy.
Another embodiment is a method for synthesizing a combinatorial library including compounds of the Formula I: 
where X is S or O;
R and R1 are individually 
where Ar is aryl; R2 is H or C1 to C6 alkyl; n is 0-3 and where the aryl moiety is optionally substituted from 1 to 7 times with any combination of H, halo, alkyl, haloalkyl, arylalkyl, alkoxy, haloalkoxy, and aralkoxy, including the step of contacting a subunit selected from the group consisting of urea and thiourea with an amine in a suitable carrier.
Yet another embodiment is composition for determining possible apoptosis induction agents for a biological substrate, comprising a combinatorial library or compounds generated therefrom.
A further embodiment of the present invention is a method of killing a cancer cell by contacting the cancer cell with a combinatorial library or compounds generated therefrom.
Another embodiment of the invention includes a kit for determining possible apoptosis induction agents for a biological substrate.
The above summary of the present invention is not intended to describe each disclosed embodiment or every implementation of the present invention. The Figures and the detailed description which follow more particularly exemplify these embodiments.